UniProt functional annotation for P08394



	UniProt functional annotation for P08394

UniProt code: P08394.

Organism: Escherichia coli (strain K12).

Taxonomy: Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales; Enterobacteriaceae; Escherichia.

Function: A helicase/nuclease that prepares dsDNA breaks (DSB) for recombinational DNA repair. Binds to DSBs and unwinds DNA via a rapid (>1 kb/second) and highly processive (>30 kb) ATP-dependent bidirectional helicase. Unwinds dsDNA until it encounters a Chi (crossover hotspot instigator, 5'-GCTGGTGG-3') sequence from the 3' direction. Cuts ssDNA a few nucleotides 3' to Chi site, by nicking one strand or switching the strand degraded (depending on the reaction conditions). The properties and activities of the enzyme are changed at Chi. The Chi-altered holoenzyme produces a long 3'-ssDNA overhang which facilitates RecA-binding to the ssDNA for homologous DNA recombination and repair. Holoenzyme degrades any linearized DNA that is unable to undergo homologous recombination (PubMed:4562392, PubMed:4552016, PubMed:123277). In the holoenzyme this subunit contributes ATPase, 3'- 5' helicase, exonuclease activity and loads RecA onto ssDNA. The RecBC complex requires the RecD subunit for nuclease activity, but can translocate along ssDNA in both directions. {ECO:0000269|PubMed:10197988, ECO:0000269|PubMed:10518611, ECO:0000269|PubMed:10766864, ECO:0000269|PubMed:123277, ECO:0000269|PubMed:12815437, ECO:0000269|PubMed:12815438, ECO:0000269|PubMed:1535156, ECO:0000269|PubMed:16041061, ECO:0000269|PubMed:1618858, ECO:0000269|PubMed:16388588, ECO:0000269|PubMed:18079176, ECO:0000269|PubMed:20852646, ECO:0000269|PubMed:23851395, ECO:0000269|PubMed:25073102, ECO:0000269|PubMed:4268693, ECO:0000269|PubMed:4552016, ECO:0000269|PubMed:4562392, ECO:0000269|PubMed:7608206, ECO:0000269|PubMed:9192629, ECO:0000269|PubMed:9230304, ECO:0000269|PubMed:9448271, ECO:0000269|PubMed:9790841}.

Catalytic activity: Reaction=Exonucleolytic cleavage (in the presence of ATP) in either 5'- to 3'- or 3'- to 5'-direction to yield 5'- phosphooligonucleotides.; EC=3.1.11.5; Evidence={ECO:0000255|HAMAP- Rule:MF_01485};

Cofactor: Name=Mg(2+); Xref=ChEBI:CHEBI:18420; Evidence={ECO:0000255|HAMAP-Rule:MF_01485, ECO:0000269|PubMed:15538360, ECO:0000269|PubMed:16388588, ECO:0000269|PubMed:18668125, ECO:0000305}; Note=Magnesium is required for both helicase and nuclease activity; its relative concentration alters helicase speed and nuclease activity in a complicated fashion. {ECO:0000255|HAMAP-Rule:MF_01485, ECO:0000269|PubMed:15538360, ECO:0000269|PubMed:16388588, ECO:0000269|PubMed:18668125, ECO:0000305};

Activity regulation: After reacting with DNA bearing a Chi site the holoenzyme is disassembled and loses exonuclease activity, DNA unwinding and Chi-directed DNA cleavage; RecB remains complexed with ssDNA, which may prevent holoenzyme reassembly (PubMed:10197988). High levels of Mg(2+) (13 mM MgCl(2+)) or incubation with DNase allow holoenyzme reassembly, suggesting it is DNA bound to RecB that prevents reassembly (PubMed:10197988). {ECO:0000269|PubMed:10197988, ECO:0000269|PubMed:1535156}.

Subunit: Heterotrimer of RecB, RecC and RecD. All subunits contribute to DNA-binding. The C-terminus interacts with RecA (PubMed:16483938). Interacts with YgbT (Cas1) (PubMed:21219465). {ECO:0000269|PubMed:15538360, ECO:0000269|PubMed:1618858, ECO:0000269|PubMed:16483938, ECO:0000269|PubMed:18668125, ECO:0000269|PubMed:21219465, ECO:0000269|PubMed:3526335, ECO:0000269|PubMed:4552016, ECO:0000269|PubMed:9448271}.

Domain: The N-terminal DNA-binding domain (residues 1-929) is a ssDNA- dependent ATPase and has ATP-dependent 3'-5' helicase function; both are stimulated in the presence of RecC, suggesting this domain interacts with RecC. Holoenzyme reconstituted with this RecB N-terminal fragment has no nuclease activity (PubMed:9448271). The C-terminal domain (residues 928-1180) has weak ssDNA endonuclease activity as an isolated fragment (PubMed:9790841) (PubMed:10518611). RecD probably interacts with this domain. The C-terminal domain interacts with RecA, facilitating its loading onto ssDNA. Interaction is decreased by ATP (PubMed:16483938). {ECO:0000269|PubMed:10518611, ECO:0000269|PubMed:16483938, ECO:0000269|PubMed:9448271, ECO:0000269|PubMed:9790841}.

Domain: The holoenzyme may undergo conformational shifts upon DNA binding: the nuclease domain of RecB may swing away from the DNA exit tunnel in RecC. When Chi DNA binds to the RecC tunnel, the nuclease domain may then swing back to its original position (that seen in crystal structures), allowing it to nick the DNA 3' of the Chi site and then rotate to load RecA. At high Mg(2+) the nuclease domain may swing back more frequently, explaining differences seen in assays performed at high Mg(2+). {ECO:0000269|PubMed:25073102}.

Disruption phenotype: Decreased degradation of DNA with free ends that is unable to undergo homologous recombination, which can fortuitously lead to more efficient viral infection (PubMed:4562392, PubMed:123277). Cells are deficient in DNA recombination repair and have increased sensitivity to UV light. The cultures have many inviable cells. {ECO:0000269|PubMed:123277, ECO:0000269|PubMed:4562392, ECO:0000269|PubMed:6389498}.

Similarity: Belongs to the helicase family. UvrD subfamily. {ECO:0000255|HAMAP-Rule:MF_01485}.

Annotations taken from UniProtKB at the EBI.


Organism:		Escherichia coli (strain K12).
Taxonomy:		Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales; Enterobacteriaceae; Escherichia.



Function:		A helicase/nuclease that prepares dsDNA breaks (DSB) for recombinational DNA repair. Binds to DSBs and unwinds DNA via a rapid (>1 kb/second) and highly processive (>30 kb) ATP-dependent bidirectional helicase. Unwinds dsDNA until it encounters a Chi (crossover hotspot instigator, 5'-GCTGGTGG-3') sequence from the 3' direction. Cuts ssDNA a few nucleotides 3' to Chi site, by nicking one strand or switching the strand degraded (depending on the reaction conditions). The properties and activities of the enzyme are changed at Chi. The Chi-altered holoenzyme produces a long 3'-ssDNA overhang which facilitates RecA-binding to the ssDNA for homologous DNA recombination and repair. Holoenzyme degrades any linearized DNA that is unable to undergo homologous recombination (PubMed:4562392, PubMed:4552016, PubMed:123277). In the holoenzyme this subunit contributes ATPase, 3'- 5' helicase, exonuclease activity and loads RecA onto ssDNA. The RecBC complex requires the RecD subunit for nuclease activity, but can translocate along ssDNA in both directions. {ECO:0000269\|PubMed:10197988, ECO:0000269\|PubMed:10518611, ECO:0000269\|PubMed:10766864, ECO:0000269\|PubMed:123277, ECO:0000269\|PubMed:12815437, ECO:0000269\|PubMed:12815438, ECO:0000269\|PubMed:1535156, ECO:0000269\|PubMed:16041061, ECO:0000269\|PubMed:1618858, ECO:0000269\|PubMed:16388588, ECO:0000269\|PubMed:18079176, ECO:0000269\|PubMed:20852646, ECO:0000269\|PubMed:23851395, ECO:0000269\|PubMed:25073102, ECO:0000269\|PubMed:4268693, ECO:0000269\|PubMed:4552016, ECO:0000269\|PubMed:4562392, ECO:0000269\|PubMed:7608206, ECO:0000269\|PubMed:9192629, ECO:0000269\|PubMed:9230304, ECO:0000269\|PubMed:9448271, ECO:0000269\|PubMed:9790841}.


Catalytic activity:		Reaction=Exonucleolytic cleavage (in the presence of ATP) in either 5'- to 3'- or 3'- to 5'-direction to yield 5'- phosphooligonucleotides.; EC=3.1.11.5; Evidence={ECO:0000255\|HAMAP- Rule:MF_01485};
Cofactor:		Name=Mg(2+); Xref=ChEBI:CHEBI:18420; Evidence={ECO:0000255\|HAMAP-Rule:MF_01485, ECO:0000269\|PubMed:15538360, ECO:0000269\|PubMed:16388588, ECO:0000269\|PubMed:18668125, ECO:0000305}; Note=Magnesium is required for both helicase and nuclease activity; its relative concentration alters helicase speed and nuclease activity in a complicated fashion. {ECO:0000255\|HAMAP-Rule:MF_01485, ECO:0000269\|PubMed:15538360, ECO:0000269\|PubMed:16388588, ECO:0000269\|PubMed:18668125, ECO:0000305};
Activity regulation:		After reacting with DNA bearing a Chi site the holoenzyme is disassembled and loses exonuclease activity, DNA unwinding and Chi-directed DNA cleavage; RecB remains complexed with ssDNA, which may prevent holoenzyme reassembly (PubMed:10197988). High levels of Mg(2+) (13 mM MgCl(2+)) or incubation with DNase allow holoenyzme reassembly, suggesting it is DNA bound to RecB that prevents reassembly (PubMed:10197988). {ECO:0000269\|PubMed:10197988, ECO:0000269\|PubMed:1535156}.
Subunit:		Heterotrimer of RecB, RecC and RecD. All subunits contribute to DNA-binding. The C-terminus interacts with RecA (PubMed:16483938). Interacts with YgbT (Cas1) (PubMed:21219465). {ECO:0000269\|PubMed:15538360, ECO:0000269\|PubMed:1618858, ECO:0000269\|PubMed:16483938, ECO:0000269\|PubMed:18668125, ECO:0000269\|PubMed:21219465, ECO:0000269\|PubMed:3526335, ECO:0000269\|PubMed:4552016, ECO:0000269\|PubMed:9448271}.
Domain:		The N-terminal DNA-binding domain (residues 1-929) is a ssDNA- dependent ATPase and has ATP-dependent 3'-5' helicase function; both are stimulated in the presence of RecC, suggesting this domain interacts with RecC. Holoenzyme reconstituted with this RecB N-terminal fragment has no nuclease activity (PubMed:9448271). The C-terminal domain (residues 928-1180) has weak ssDNA endonuclease activity as an isolated fragment (PubMed:9790841) (PubMed:10518611). RecD probably interacts with this domain. The C-terminal domain interacts with RecA, facilitating its loading onto ssDNA. Interaction is decreased by ATP (PubMed:16483938). {ECO:0000269\|PubMed:10518611, ECO:0000269\|PubMed:16483938, ECO:0000269\|PubMed:9448271, ECO:0000269\|PubMed:9790841}.
Domain:		The holoenzyme may undergo conformational shifts upon DNA binding: the nuclease domain of RecB may swing away from the DNA exit tunnel in RecC. When Chi DNA binds to the RecC tunnel, the nuclease domain may then swing back to its original position (that seen in crystal structures), allowing it to nick the DNA 3' of the Chi site and then rotate to load RecA. At high Mg(2+) the nuclease domain may swing back more frequently, explaining differences seen in assays performed at high Mg(2+). {ECO:0000269\|PubMed:25073102}.
Disruption phenotype:		Decreased degradation of DNA with free ends that is unable to undergo homologous recombination, which can fortuitously lead to more efficient viral infection (PubMed:4562392, PubMed:123277). Cells are deficient in DNA recombination repair and have increased sensitivity to UV light. The cultures have many inviable cells. {ECO:0000269\|PubMed:123277, ECO:0000269\|PubMed:4562392, ECO:0000269\|PubMed:6389498}.
Similarity:		Belongs to the helicase family. UvrD subfamily. {ECO:0000255\|HAMAP-Rule:MF_01485}.